Macrophages and synovial fibroblasts in the joint lining are central to the destruction observed in rheumatoid arthritis (RA). Although Fas and Fas ligand (FasL) are expressed on cells in the synovial lining, in vivo apoptosis is rarely observed. We have demonstrated that Flice Like Inhibitory Protein (Flip) is highly expressed in the RA joint. Suppression of the expression of Flip resulted in Fas-mediated apoptosis of normal macrophages and those from the RA joint. Our preliminary data demonstrates that following the interruption of Fas/FasL interactions on macrophages, activation of NF-kappaB and the expression of IL-6, mediated through the toll like receptor (TLR) and IL-1 receptor 1 (IL-1R1) pathways, was suppressed. Further, experiments in RAW 264.7 macrophage-like cells demonstrate that the ectopic expression of either Flip or FADD, which are recruited to the Fas Death Inducing Signal Complex (DISC) following Fas ligation, suppressed the LPS-induced activation of NF-kappaB and IL-6 promoter reporters. In 293 cells, which lack TLRs, neither Flip or FADD suppressed TNFalpha-induced NF-kappaB or IL-6 promoter activation. These observations suggest that following Fas/FasL interaction, as occurs in the lining in the RA joint, Flip and FADD are recruited to the Fas DISC, promoting enhanced activation through the TLR/IL-1 R1 pathway. Although both isoforms of Flip (FlipL and FlipS) are capable of protecting against apoptosis and modulating the TLR/IL-1R1 pathway, Flips is uniquely expressed in macrophagess, compared to other cell types such as synovial fibroblasts. Our preliminary data suggests that FlipL and FlipS are differentially regulated at the post-transcriptional level in a cell type-specific fashion. We propose in aim 1 to examine the mechanisms responsible for the modulation of activation through the TLR/IL-1 R1 pathway. Aim 2 proposes to characterize the mechanisms responsible for regulating the cell type-specific expression of the Flip isoforms. Since the in vivo effects of deletion of flip have not been examined, aim 3 proposes to examine the effects of the cell type-specific deletion of flip in macrophages in mice. These studies will provide insights into mechanisms that contribute to the pathogenesis of RA and potentially the development of new therapeutic strategies.